昆虫学报 ›› 2022, Vol. 65 ›› Issue (3): 271-279.doi: 10.16380/j.kcxb.2022.03.003

• 研究论文 • 上一篇    下一篇

草地贪夜蛾三个SfruPBPs对本种及同域种劳氏粘虫性信息素及腺体组分的亲和力

郑树壕1, 司玉晓1, 闫祺1, 郭慧芳2, 董双林1,*   

  1.  (1. 南京农业大学植物保护学院, 农作物生物灾害综合治理教育部重点实验室, 南京 210095; 2. 江苏省农业科学院植物保护研究所, 南京 210014)
  • 出版日期:2022-03-20 发布日期:2022-03-24

Binding affinities of three SfruPBPs to sex pheromone and gland components from Spodopetera frugiperda and sympatric Leucania loreyi (Lepidoptera: Noctuidae)

ZHENG Shu-Hao1, SI Yu-Xiao1, YAN Qi1, GUO Hui-Fang2, DONG Shuang-Lin1,*   

  1. (1. Key Laboratory of Integrated Management of Crop Diseases and Pests, Ministry of Education, College of Plant Protection, Nanjing Agricultural University, Nanjing 210095, China; 2. Institute of Plant Protection, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, China)
  • Online:2022-03-20 Published:2022-03-24

摘要:

【目的】测定草地贪夜蛾Spodoptera frugiperda 3个信息素结合蛋白(pheromone binding protein, PBP)(SfruPBP)对草地贪夜蛾及同域近缘种劳氏粘虫Leucania loreyi性信息素及腺体组分的结合特性,探究草地贪夜蛾这3个SfruPBPs在两种昆虫不同性信息素组分识别中的作用。【方法】以pET-30a(+)为载体分别构建草地贪夜蛾3个SfruPBPs的原核表达质粒,导入大肠杆菌Escherichia coli并培养至OD600=0.6~0.8后,加入IPTG进行这3个SfruPBPs的诱导表达,采用Ni-NTA琼脂糖磁珠进行蛋白纯化;利用荧光竞争结合实验测定这3个SfruPBPs与草地贪夜蛾和劳氏粘虫的共14种性信息素和腺体组分的结合特性。【结果】原核表达获得SfruPBP1-3重组蛋白,进一步纯化后电泳检测得到单一且预期大小的目的蛋白。荧光竞争结合实验测定发现,在草地贪夜蛾和劳氏粘虫已报道的总计14种性信息素和腺体组分中,SfruPBP1对这两种害虫的性信息素主组分Z9-14∶Ac特异性强结合,Ki=0.80 μmol/L;SfruPBP2结合谱较广,除对性信息素次要组分Z7-12∶Ac及腺体组分12∶Ac, E7-12∶Ac和Z10-14∶Ac强结合(Ki<1.00 μmol/L)外,还对性信息素主组分Z9-14∶Ac及腺体组分Z9-12∶Ac和11-12∶Ac具中等结合能力(1.00 μmol/L<Ki<4.00 μmol/L);SfruPBP3仅对腺体组分Z912∶Ac具中等结合能力(Ki=1.36 μmol/L)。在上述与SfruPBP具有强或中等结合能力的组分中,Z9-14∶Ac, Z7-12∶Ac及12∶Ac也是劳氏粘虫的性信息素或腺体组分,但3个SfruPBPs对劳氏粘虫的特有腺体组分Z7-14∶Ac均无明显结合能力。【结论】草地贪夜蛾3个SfruPBPs均在其性信息素感受中发挥作用,但不同SfruPBPs具有明显的组分选择性;这3个SfruPBPs对劳氏粘虫的腺体特有组分Z7-14∶Ac均无明显结合能力。

关键词: 草地贪夜蛾, 劳氏粘虫, 信息素结合蛋白, 原核表达, 荧光竞争结合

Abstract: 【Aim】 This study aims to determine the binding characteristics of three pheromone binding proteins (PBPs) from the fall armyworm, Spodoptera frugiperda with sex pheromone and gland components of S. frugiperda and sympatric Leucania loreyi, and thus to investigate the roles of the three SfruPBPs in the recognition of pheromone components of these two species. 【Methods】 Plasmids were constructed using pET-30a(+) as the prokaryotic expression vector of the three SfruPBPs of S. frugiperda and introduced into Escherichia coli. After the OD600 value of the culture of E. coli reached 0.6-0.8, the three SfruPBPs were induced to express using IPTG, and purified by the Ni-NTA magnetic agarose beads. The binding characteristics of the three SfruPBPs with 14 sex pheromone and gland components of S. frugiperda and L. loreyi were assayed by fluorescence competitive binding assay. 【Results】 The recombinant PBP proteins SfruPBP1-3 were obtained by prokaryotic expression. Single target protein bands of expected size were obtained in the agarose gel electrophoresis after protein purification. Fluorescence competitive binding assay revealed that among the reported 14 sex pheromone and gland components of S. frugiperda and L. loreyi, SfruPBP1 had strong specific binding ability with the major pheromone component Z9-14∶Ac (Ki=0.80 μmol/L) of these two pest species, and SfruPBP2 displayed broad ligand spectrum, showing strong binding ability with the minor pheromone component Z7-12∶Ac and the gland components 12∶Ac, E7-12∶Ac and Z10-14∶Ac (Ki<1.00 μmol/L), and medium binding ability with the major pheromone component Z9-14∶Ac and the gland components Z9-12∶Ac and 11-12∶Ac (1.00 μmol/L<Ki<4.00 μmol/L). SfruPBP3 had medium binding ability only with the gland component Z9-12∶Ac, with the Ki value of 1.36 μmol/L. Among the chemicals with strong or medium binding affinities to SfruPBP, Z9-14∶Ac, Z7-12∶Ac and 12∶Ac were also sex pheromone or gland components of L. loreyi, however, all the three SfruPBPs showed no obvious binding ability with Z7-14∶Ac, a specific gland component in L. loreyi. 【Conclusion】 All the three SfruPBPs of S. frugiperda play important roles in the sex pheromone perception, but each SfruPBP displays a distinct selectivity to components. The three SfruPBPs show no obvious binding ability with the specific gland component Z7-14∶Ac of L. loreyi.

Key words: Spodoptera frugiperda, Leucania loreyi, pheromone binding protein, prokaryotic expression, fluorescence competitive binding